POS1044 SINGLE CELL TRANSCRIPTOMICS ANALYSIS OF BLOOD IMMUNE CELLS IDENTIFIES GENE SIGNATURES AND CELL SUBSETS ASSOCIATED WITH DISEASE ACTIVITY IN A DIVERSE POPULATION OF RHEUMATOID ARTHRITIS PATIENTS
Authors: M. Binvignat, B. Miao, C. Wibrand, M. Yang, D. Rychkov, E. Flynn, J. Nititham, A. Carvidi, M. Krueger, E. Niemi, Y. Sun, E. Mariotti-Ferrandiz, J. Sellam, C. Ye, A. Butte, L. Criswell, M. Nakamura, M. Sirota
Journal: Annals of the Rheumatic Diseases (2023)

<h3>Background</h3> Single cell transcriptional profiling has previously been used to identify gene signatures associated with rheumatoid arthritis (RA) pathophysiology, as well as characterize cell subpopulations. However previous studies have not adequately addressed potential comparison bias such as age and sex, or included patients from different ethnic backgrounds. <h3>Objectives</h3> This study leverages single cell sequencing of peripheral blood mononuclear cells (PBMCs) from patients with RA and matched healthy controls to identify disease relevant cell subsets and cell type specific signatures of disease in the periphery across a diverse cohort of patients. <h3>Methods</h3> PBMC samples from 18 RA patients and 18 healthy controls matched on age, sex, and ethnicity were sequenced at a single cell resolution using 10X Chromium protocols. Samples were processed using standard CellRanger and Scanpy pipelines, with HarmonyPy for batch correction. Differential gene expression was computed using DESeq2 and pathway analysis was carried out with GSEA. Cell proportion analyses were carried out within cell subsets using Mann-Whitney analysis between matched RA and control and in RA remission and low activity (DAS28 CRP &lt; 3.2) versus moderate and high (DAS28 CRP ≥ 3.2). <h3>Results</h3> The final dataset consisted of 22,159 genes across 125,698 cells. The cohort was composed of 66.7% of females with a mean age of 54 years old, and was multi-ethnic with 11.1% of asian-american, 83.5 % of caucasian-american and 5.6 % of hispanic american, the mean DAS28-CRP for patients with RA was 3.33, 61.1 % had positive rheumatoid factor and 83.3 % had anti-citrullinated protein antibodies, 38.9 % were treated by DMARDs, only one patient had a biological. We identified 18 peripheral blood mononuclear cells (PBMC) subsets, including 5 CD4+ T cell subsets, 3 CD8+ T cell subsets, 2 Natural Killer (NK) cell subsets, 3 B cell subsets, and 5 monocyte subsets (Figure 1A). 70 genes were found to be significantly differentially expressed between RA and control CD4+ T cells, 98 genes in monocytes, 11 in B cells, 97 in CD8+ T cells, and 23 in NK cells (FDR ≤ 0.05) with some overlapping across cell types (Figure 1B). Enrichment analysis of genes significantly upregulated in RA-derived classical monocytes identified pathways associated with type 1 interferon signaling and cytokine production (p&lt;0.001). Density plots revealed shifts in proportions of cell subsets between patients with high and moderate activity (DAS28 CRP ≥ 3.2;n=7), patients in low disease activity and remission (DAS28 CRP &lt; 3.2; n=9), and healthy controls (n=18, Figure 1C). Non classical monocytes were significantly decreased in patients with low disease activity compared to control and high disease activity (p = 0.015 and 0.022). CD4 T central memory cells were also decreased in patients with high disease activity (p = 0.034) compared with controls. <h3>Conclusion</h3> Here we describe a dataset of single cell RNA sequenced PBMCs from a diverse population of patients with RA and matched healthy controls. We identified differentially expressed genes and new cell subsets linked to RA but also disease activity, providing potential new insight in non-remission mechanisms and pathophysiology in a diverse RA cohort. <h3>References</h3> [1] Zhang F et al. Nat Immunol. 2019 PMID: 31061532 [2] Wu X et al. Nat Commun. 2021 PMID: 34404786 <h3>Acknowledgements:</h3> NIL. <h3>Disclosure of Interests</h3> None Declared.